Assay design

What does "Flex-Seq assay design" mean?

There are multiple SNPs within 100 nt of my SNP of interest. Will the presence of these surrounding SNPs significantly impact oligonucleotide probe design?

What parameters do we use to sequence SNPs?

How are the parameters set to account for statistical significance or reliability of the SNP call when dealing with possible sequencing errors?

Can Flex-Seq be used to detect off-target SNPs with CRISPR-Cas9, based on targeting regions of binding site homology (or similar)?

What factors can cause a Flex-Seq probe design to fail?

I have already performed KASP on my samples. Can I use my KASP assays directly for a Flex-Seq project?

I have a selection of SNPs I wish to test via Flex-Seq, but I am unsure how successful they will be in the design process. Will I be able to exchange my SNPs prior to library generation?

Can Flex-Seq be used with SNPs (single nucleotide polymorphisms), InDels (insertions and deletions) and repeats?

What is the expected read length?

What is the expected read coverage (depth)?

Will LGC, Biosearch Technologies be able to provide any service or guidance for pre-submission bioinformatics, for example, assembly of the reference sequence or advice on which selection of markers would be most appropriate?

I do not know the exact make-up of my SNPs, but I know their genomic location. Is this information sufficient for Flex-Seq?

Can reference scaffolds be used in place of reference chromosome when submitting reference sequences?

I do not have genomic sequence for my particular organism. Can I use a closely related organism/species for my reference sequences?

For the reference genome, will the name of reference sequence in a publically available database be sufficient (e.g. <a href="https://www.ncbi.nlm.nih.gov/genome/51?genome_assembly_id=322645" target="_blank">GRCh38</a> from NCBI)?

My reference genome was assembled from fragment DNA sequencing, and is highly polymorphic (with possible unidentified SNPs in the flanking regions to my SNPs of interest). Would Flex-Seq work?

Will a score be given to each designed probe to estimate its success prior to sequencing so I can select the desired SNPs for library generation?

I have determined how many markers I wish to characterise using Flex-Seq. What is the recommended number of candidates to ensure successful probe design across all my markers?

Question?